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•31 neurochemicals-targeted profiling could be used in CNS drug discovery.•The neurochemical profiling represents comprehensive brain neurochemistry.•Nefazodone modulates cholinergic, ...serotonergic, GABAergic, and dopaminergic systems.
Depression, typically understood as a mental disorder, is characterized by monoamine neurotransmitter deficiencies in the central nervous system (CNS). A majority of antidepressants have been developed to increase endogenous levels of monoamine neurotransmitters, such as dopamine (DA), norepinephrine (NE), and serotonin (5-HT) in brain tissue. Increasingly, the critical effects of a variety of neurotransmitters, including those of the dopaminergic, serotonergic, kynurenergic, cholinergic, histaminergic, and GABAergic systems, as well as several amino acids, have been spotlighted as neuromodulators over the years in various behavioral and emotional responses. In the present study, we developed a simple method for simultaneous analysis of 31 neurochemicals in mouse whole brain tissue using liquid chromatography tandem mass spectrometry (LC-MS/MS). Our neurochemical profiles were classified into seven categories according to metabolic pathways composed of neurotransmitters and their precursors or metabolites. The analytical method was validated by intra- and inter-day precision, accuracy, sensitivity, and selectivity, and finally applied for neurochemical profiling in the mouse brain following nefazodone treatment. In this study, the modulation of 5-HT and DA by nefazodone was confirmed in the mouse brain, as previously reported. Interestingly, nefazodone-induced broad-spectrum neurochemical changes could suggest that nefazodone has novel anti-depressive and anti-cholinesterase potency via increasing endogenous levels of betaine and acetylcholine in the mouse brain. This neurochemical profiling approach may represent a meaningful tool for better understanding brain neurochemistry and neurotoxicity in discovery and development of CNS drugs.
Tanshinone IIA is a diterpene quinone isolated from the roots of
bunge that has traditionally been used in China for the treatment of cardiovascular and cerebrovascular disorders. Although there is ...recent evidence showing that tanshinone IIA has an anti-obesity effect, its underlying mechanism of anti-obesity effect is poorly understood. Here, we investigated the effect of tanshinone IIA on lipid accumulation in 3T3-L1 preadipocytes and zebrafish. Notably, tanshinone IIA at 10 μM concentration greatly reduced lipid accumulation and triglyceride (TG) contents during 3T3-L1 preadipocyte differentiation, suggesting its anti-adipogenic effect. On mechanistic levels, tanshinone IIA reduced the expression levels of CCAAT/enhancer-binding protein-α (C/EBP-α), peroxisome proliferator-activated receptor-γ (PPAR-γ), fatty acid synthase (FAS), and perilipin A but also the phosphorylation levels of signal transducer and activator of transcription-3/5 (STAT-3/5) in differentiating 3T3-L1 cells. In addition, tanshinone IIA strongly inhibited leptin and resistin mRNA expression in differentiating 3T3-L1 cells. Importantly, the tanshinone IIA's lipid-reducing effect was also seen in zebrafish. In sum, these findings demonstrate that tanshinone IIA has anti-adipogenic effects on 3T3-L1 cells and zebrafish, and its anti-adipogenic effect on 3T3-L1 cells is largely attributable to the reduced expression and/or phosphorylation levels of C/EBP-α, PPAR-γ, FAS, perilipin A, and STAT-3/5.
A macrocyclic peptide scaffold with well-established structure–property relationship is desirable for tackling undruggable targets. Here, we adopted a natural macrocycle, cyclosporin O (CsO) and its ...derivatives (CP1–3), and evaluated the impact of conformation on membrane permeability, cyclophilin A (CypA) binding, and the pharmacokinetic (PK) profile. In nonpolar media, CsO showed a similar conformation to cyclosporin A (CsA), a well-known chameleonic macrocycle, but less chameleonic behavior in a polar environment. The weak chameleonicity of CsO resulted in decreased membrane permeability; however, the more rigid conformation of CsO was not detrimental to its PK profile. CsO exhibited a higher plasma concentration than CsA, which resulted from minimal CypA binding and lower accumulation in red blood cells and moderate oral bioavailability (F = 12%). Our study aids understanding of CsO, a macrocyclic peptide that is less explored than CsA but with greater potential for diversity generation and rational design.
Recently, with the development of computer vision using artificial intelligence (AI), clinical research on diagnosis and prediction using medical image data has increased. In this study, we applied ...AI methods to analyze hepatic fibrosis in mice to determine whether an AI algorithm can be used to analyze lesions. Whole slide image (WSI) Sirius Red staining was used to examine hepatic fibrosis. The Xception network, an AI algorithm, was used to train normal and fibrotic lesion identification. We compared the results from two analyses, that is, pathologists’ grades and researchers’ annotations, to observe whether the automated algorithm can support toxicological pathologists efficiently as a new apparatus. The accuracies of the trained model computed from the training and validation datasets were greater than 99%, and that obtained by testing the model was 100%. In the comparison between analyses, all analyses showed significant differences in the results for each group. Furthermore, both normalized fibrosis grades inferred from the trained model annotated the fibrosis area, and the grades assigned by the pathologists showed significant correlations. Notably, the deep learning algorithm derived the highest correlation with the pathologists’ average grade. Owing to the correlation outcomes, we conclude that the trained model might produce results comparable to those of the pathologists’ grading of the Sirius Red-stained WSI fibrosis. This study illustrates that the deep learning algorithm can potentially be used for analyzing fibrotic lesions in combination with Sirius Red-stained WSIs as a second opinion tool in non-clinical research.
Microplastics, small pieces of plastic derived from polystyrene, have recently become an ecological hazard due to their toxicity and widespread occurrence in aquatic ecosystems. In this study, we ...exposed zebrafish larvae to two types of fluorescent polystyrene nanoparticles (PS-NPs) to identify their size-dependent effects. PS-NPs of 50 nm, unlike 100 nm PS-NPs, were found to circulate in the blood vessels and accumulate in the brains of zebrafish larvae. Behavioral and electroencephalogram (EEG) analysis showed that 50 nm PS-NPs induce abnormal behavioral patterns and changes in EEG power spectral densities in zebrafish larvae. In addition, the quantification of endogenous neurochemicals in zebrafish larvae showed that 50 nm PS-NPs disturb dopaminergic metabolites, whereas 100 nm PS-NPs do not. Finally, we assessed the effect of PS-NPs on the permeability of the blood–brain barrier (BBB) using a microfluidic system. The results revealed that 50 nm PS-NPs have high BBB penetration compared with 100 nm PS-NPs. Taken together, we concluded that small nanoparticles disturb the nervous system, especially dopaminergic metabolites.
Background. Cuban sugarcane wax acids (SCWA) and policosanol (PCO) are mixtures of higher aliphatic acids and alcohols, respectively, purified from sugarcane wax with different chief components. ...Although it has been known that they have antioxidant and anti-inflammatory activities, physiological properties on molecular mechanism of SCWA have been less studied than PCO. Methods. In this study, we compared antiatherogenic activities of SCWA and PCO via encapsulation with reconstituted high-density lipoproteins (rHDL). Results. After reconstitution, SCWA-rHDL showed smaller particle size than PCO-rHDL with increase of content. PCO-rHDL or SCWA-rHDL showed distinct inhibition of glycation with similar extent in the presence of fructose. PCO-rHDL or SCWA-rHDL showed strong antioxidant activity against cupric ion-mediated oxidation of low-density lipoproteins (LDL), and inhibition of oxLDL uptake into macrophages. Although PCO-rHDL showed 1.2-fold stronger inhibition against cholesteryl ester transfer protein (CETP) activity than SCWA-rHDL, SCWA-rHDL enhanced 15% more brain cell (BV-2) growth and 23% more regeneration of tail fin in zebrafish. Conclusion. PCO and SCWA both enhance the beneficial functions of HDL to maximize its antioxidant, antiglycation, and antiatherosclerotic activities and the inhibition of CETP. These enhancements of HDL functionality by PCO and SCWA could exert antiaging and rejuvenation activity.
The current study was designed to investigate the short-term effects of policosanol consumption on blood pressure (BP) and the lipid parameters in healthy Korean participants with prehypertension. A ...total of 84 healthy participants were randomly allocated to three groups receiving placebo, 10 mg of policosanol, or 20 mg of policosanol for 12 weeks. Based on an average of three measurements of peripheral BP, the policosanol 20 mg group exhibited the most significant reduction, that is, up to 7.7% reduction of average systolic BP (SBP) from 136.3 ± 6.1 mmHg (week 0) to 125.9 ± 8.6 mmHg (week 12,
< 0.001). Between group comparisons using repeated measures ANOVA showed that the policosanol 20 mg group had a significant reduction of SBP at 12 weeks (
= 0.020) and a reduction of diastolic BP (DBP) at 8 weeks (
= 0.041) and 12 weeks (
= 0.035). The policosanol 10 mg and 20 mg groups showed significant reductions in aortic SBP of 7.4% and 8.3%, respectively. The policosanol groups showed significant reductions of total cholesterol (TC) of 9.6% and 8.6% and low-density lipoproteins (LDL-C) of 21% and 18% for 10 mg and 20 mg of policosanol, respectively. Between group comparisons using repeated measures ANOVA showed that the policosanol (10 mg and 20 mg) groups at 12 weeks had a significant reduction of TC (
= 0.0004 and
= 0.001) and LDL-C (
= 0.00005 and
= 0.0001) and elevation of %HDL-C (
= 0.048 and
= 0.014). In conclusion, 12-week consumption of policosanol resulted in significant reductions of peripheral SBP and DBP, aortic SBP and DBP, mean arterial pressure (MAP), and serum TC and LDL-C with elevation of % HDL-C.
Emerging evidence indicates that NADPH oxidase (NOX) and its reactive oxygen species (ROS) products modulate a variety of cellular events, including proliferation, differentiation, and apoptosis. In ...this study, we investigated the functions of NOX2 and ROS in immune modulation using NOX2 knockout (KO) mice. Interestingly, NOX2 KO mice spontaneously developed arthritis with onset at 6-7 wk of age and high incidence (60%) at 15-18 wk of age. Arthritis severity in NOX2 KO mice was proportionally increased with age and higher in females than in males. Bone destruction was confirmed by microcomputed tomography scanning and histological analyses of joints. Inflammatory factors, including TNF-α, IL-1β, and RANKL, and serum level of anti-type II collagen IgG were significantly increased in NOX2 KO mice. In addition, NOX2 deficiency perturbed the immune system upon aging. NOX2 KO mice demonstrated preferred development of CD11b+Gr-1+ myeloid cells with profound production of proinflammatory cytokines and augmented expression of IL-17 through the activation of STAT3 and RORγt in vivo. NOX2 deficiency increased differentiation of effector Th cells in vitro and decreased CD25+FoxP3+ Treg cells both in vitro and in vivo. Furthermore, adoptive transfer of NOX2-deficient CD4⺠T cells into RAG KO mice increased arthritic inflammation compared with WT cells. These results demonstrated that NOX2 deficiency affected the development of CD11b+ myeloid cells and Th17/Treg cells, and thus promoted inflammatory cytokine production and inflammatory arthritis development, strongly supporting a crucial role for ROS generation in the modulation of Th17/Treg cell development and its related inflammatory immune response upon aging.
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PPARγ agonists induced obesity in animal models as a side effect. Microarray experiments reveal that PPARγ agonist upregulates the expression of lipin-1 and this upregulation is ...correlated with the activity of the agonists. Lipin-1 induced by PPARγ agonists decreased the levels of PPARγ and ERK1/2 phosphorylation through direct interaction with these proteins in 3T3-L1 cells. In PPARγ agonist-treated 3T3-L1 preadipocytes, the knockdown of lipin-1 expression by small interfering RNA inhibited the adipogenesis that was induced by PPARγ agonists. In contrast, PPARγ2 expression was increased, and lipid droplets were accumulated in lipin-1-overexpressing 3T3-L1 adipocytes. Rosiglitazone (RGZ), a strong PPARγ agonist, synergistically promoted PPARγ dephosphorylation and adipogenesis in lipin-1-overexpressing 3T3-L1 preadipocytes. Therefore, lipin-1 has dual functions as a transcriptional cofactor and phosphatidate phosphatase (PAP) in the differentiation of preadipocyte cells induced by strong PPARγ agonists. In addition, the adipogenesis of 3T3-L1 cells was markedly upregulated by diacylglycerol (DAG), which was produced by lipin-1. Therefore, lipin-1 induction by PPARγ agonists might be an important factor in understanding the biological mechanism of the agonists’ adverse effects, and this information may be valuable in the development of type-2 diabetes mellitus (T2DM) therapeutics with reduced adverse effects and greater tolerability.
Abstract
Metabolic Dysfunction-Associated Steatotic Liver Disease (MASLD) is currently the leading cause of chronic liver disease worldwide. Metabolic Dysfunction-Associated Steatohepatitis (MASH), ...an advanced form of MASLD, can progress to liver fibrosis, cirrhosis, and hepatocellular carcinoma. Based on recent findings by our team that liver 5HT
2A
knockout male mice suppressed steatosis and reduced fibrosis-related gene expression, we developed a peripheral 5HT
2A
antagonist, compound
11c
for MASH. It shows good in vitro activity, stability, and in vivo pharmacokinetics (PK) in rats and dogs. Compound
11c
also shows good in vivo efficacy in a diet-induced obesity (DIO) male mice model and in a choline-deficient, L-amino acid-defined, high-fat diet (CDAHFD) male mice model, effectively improving histologic features of MASH and fibrosis. According to the tissue distribution study using
14
C-labeled
11c
, the compound was determined to be a peripheral 5HT
2A
antagonist. Collectively, first-in-class compound
11c
shows promise as a therapeutic agent for the treatment of MASLD and MASH.
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